ABSTRACT Borrelia mayonii is the causative agent of a newly-recognized tick-borne disease. First reported in the upper Midwest during February, 2016, the geographic range of this emerging public health threat is not yet known. Although B. mayonii is related to the classic Lyme disease agent, Borrelia burgdorferi, the two species exhibit substantial genetic differences. We hypothesize that those differences are responsible for the distinct features of B. mayonii infection. A striking difference between the two diseases is the high spirochetemia that has been observed only in patients infected with B. mayonii. Other symptoms of B. mayonii infection include macular rashes across the body, myalgia, and neurological signs, with some patients requiring hospitalization. Three critical questions about B. mayonii and its associated disease are being addressed by our studies: (1) What are the extents of spirochetemia, tissue pathology, and other infectious qualities in untreated mammalian B. mayonii infection? (2) What is responsible for the high-level spirochetemia that occurs during B. mayonii infection, but not during B. burgdorferi infection? and (3) What antigenic proteins are produced during B. mayonii infection, but not during B. burgdorferi infection, that can be used for diagnosis of this emerging disease? Results from our studies will provide valuable insight on the pathogenic properties of B. mayonii and lay important groundwork for diagnosis and treatment.